MHC class II combined immunodeficiency (CID or bare lymphocyte syndrome (BLS)) is an autosomal recessive congenital disease whose hallmark is the absence of major histocompatibility complex (MHC) class II proteins on all tissues. Patients with class II CID usually die in childhood of overwhelming viral and bacterial infections unless they undergo bone marrow transplantation. The defect in patients with CID has been attributed to a trans-acting factor(s) involved in the transcriptional regulation of class II gene expression since l) no mutations have been found in the class Il genes themselves, 2) the MHC locus and the disease loci segregate independently, and 3) class II expression in B cells from CID patients can be restored by somatic cell fusion to class II(+) B cells. Complementation analysis between cell lines derived from CID patients reveals that mutations in at least four different genes lead to an MHC class Il deficient phenotype. A cloning strategy utilizing a mammalian Epstein-Barr Virus (EBV) based expression system coupled with a powerful immunoselection step has recently been used to successfully isolate the group II CID gene, termed CIITA, from a cell line RJ2.2.5 in this group. The specific aims of this application focus on the analysis of the function and mechanism of action of the CIITA gene and also include the further use of this powerful cloning strategy to isolate the gene defective in two patients who form a new group, Group V, we have just identified. We propose to: l) determine the mechanism by which the CIITA protein serves as a co-activator and controller of class II gene transcription by identifying the DNA-binding proteins with which it interacts; 2) identify the mutation in the CIITA gene of clone 13, an additional member of complementation group Il whose in vivo genomic footprint and pattern of class Il expression differs from that of the other members of this group and to re-create its unique phenotype of isotype-specific defects in class Il expression in vivo by performing a targeted replacement of the wild-type CIITA gene with the clone 13 mutant gene; 3) isolate the gene responsible for MHC class II deficiency in group V CID whose phenotype suggests cell type specific expression of class Il and determine the constitutive and cytokine-inducible tissue distribution of RNA and protein for this gene using the cloned cDNA and antibodies reactive with the encoded protein as tools. Isolation and characterization of these genes should provide insight into the molecular mechanisms of human class Il gene transcription as well as the specific mutations in individual patients with CID and provide therapeutic options for isotype- specific, cell type specific control of class II expression in autoimmune disorders.